The invention relates to a method of folding recombinant somatotropins. Somatotropins are hormones which are secreted by the adenohypophysis (anterior lobe of the pituitary gland) and are known to affect the rate of skeletal growth and gain in body weight. Administration of somatotropin has been shown to cause an increase in milk production in lactating animals such as dairy cows and goats. With the advent of recombinant DNA technology, the ability to produce greater quantities of proteins, like somatotropins, in transformed microorganisms has been achieved. Nevertheless, greater amounts of protein production in some bacteria leads to bioinactive cytoplasmic aggregates containing partially folded and unfolded proteins, or both, typically referred to as inclusion bodies, or refractile bodies. For the many proteins that contain disulfide crosslinks, this problem is especially severe since bacteria used as recombinant hosts like the commonly used Escherichia coli, are not adapted to the process of sulfhydryl oxidation to the disulfide form.
The proper conformation of a protein is essential for biological activity. Therefore, protocols are developed to enable the partially folded and unfolded proteins, typically found in refractile bodies to properly fold into their biologically active conformation. Known procedures for folding proteins include the use of detergents like sodium dodecyl sulfate (SDS) as denaturing agents, see Rausch et al., U.S. Pat. No. 4,677,196. Seely, James E., U.S. Pat. No. 4,656,255, describes a process for recovering proteins that includes resolubilization of sidestream precipitates isolated from chromatography effluent. The use of an aqueous urea solution to solubilize refractile bodies containing somatotropin protein and naturation of the proteins in the urea solution was described in Bentle et al., U.S. Pat. No. 4,652,630. However, many of the known procedures result in slow folding rates, low yield and uneconomical operations. As demonstrated by this invention, it has been discovered that the use of 2-amino-2-methyl-1-propanol (AMP) can enhance the folding rate without sacrificing yields.